40 results about "Splice Donor Site" patented technology

Disclosed are materials and methods for treating diseases of the mammalian eye, and in particular, Usher syndrome 1B (USH1B). The invention provides AAV-based, dual-vector systems that facilitate the expression of full-length proteins whose coding sequences exceed that of the polynucleotide packaging capacity of an individual AAV vector. In one embodiment, vector systems are provided that include i) a first AAV vector polynucleotide that includes an inverted terminal repeat at each end of the polynucleotide and a suitable promoter followed by a partial coding sequence that encodes an N-terminal portion of a full-length polypeptide; and ii) a second AAV vector polynucleotide that includes an inverted terminal repeat at each end of the polynucleotide and a partial coding sequence that encodes a C-terminal portion of a full-length polypeptide, optionally followed by a polyadenylation (pA) signal sequence. In another embodiment, the vector system includes i) a first AAV vector polynucleotide comprising an inverted terminal repeat at each end, a suitable promoter followed by a partial coding sequence that encodes an N-terminal portion of a full-length polypeptide followed by a splice donor site and intron and ii) a second AAV vector polynucleotide comprising an inverted terminal repeat at each end, followed by an intron and a splice-acceptor site for the intron, followed by a partial coding sequence that encodes a C-terminal portion of a full-length polypeptide, optionally followed by a polyadenylation (pA) signal sequence. The coding sequence or the intron sequence in the first and second AAV vectors preferably includes a sequence region that overlaps.

The invention provides a viral self-inactivating (SIN) vector on the basis of the avian sarcoma leukosis virus (ASLV) as well as a split-packaging system comprising in addition to the SIN vector a first helper plasmid serving for the expression of the viral fusion protein gag-pol and a second helper plasmid serving for the expression of the retroviral envelope protein (env). The first and second helper plasmid, for example contained in a packaging cell line or transiently transfected, serve for the generation of non-replicating (RCR-incompetent) viral particles containing RNA having a SIN LTR according to the invention at the 3′ terminus, wherein the RNA can have a therapeutically effective section which e.g. is denoted a transgene. This 3′ SIN LTR contains an extensive deletion of the U3 region which in the course of the reverse transcription is copied into the 5′ LTR. In addition, in the SIN vector all coding regions of ASLV as well as the retroviral splice donor site are removed.